TFEB (transcription factor EB) and TFE3 (transcription factor E3) are “master regulators” of autophagy and lysosomal biogenesis. The stress response p38 mitogen-activated protein (MAP) kinases affect multiple intracellular responses including inflammation, cell growth, differentiation, cell dying, senescence, tumorigenesis, and autophagy. Small molecule p38 MAP kinase inhibitors for instance SB202190 are broadly found in dissection of related signal transduction mechanisms including redox biology and autophagy. Here, we initially aimed to check out the hyperlinks between p38 MAP kinase and TFEB/TFE3-mediated autophagy and lysosomal biogenesis. All of a sudden, we learned that only SB202190, rather of other p38 inhibitors, promotes TFEB and TFE3 to translocate within the cytosol to the nucleus and subsequently enhances autophagy and lysosomal biogenesis. Furthermore, siRNA-mediated Tfeb and Tfe3 knockdown effectively attenuated SB202190-caused gene expression and lysosomal biogenesis. Mechanistical studies shown that TFEB and TFE3 activation because of SB202190 depends on PPP3/calcineurin rather close to the inhibition of p38 or MTOR signaling, the main path for controlling TFEB and TFE3 activation. Importantly, SB202190 elevated intracellular calcium levels, and calcium chelator BAPTAP-AM blocked SB202190-caused TFEB and TFE3 activation additionally to autophagy and lysosomal biogenesis. In addition, endoplasmic reticulum (ER) calcium is required for TFEB and TFE3 activation because of SB202190. To conclude, we identified a formerly uncharacterized role of SB202190 in activating TFEB- and TFE3-dependent autophagy and lysosomal biogenesis via ER calcium release and subsequent calcium-dependent PPP3/calcineurin activation, leading to dephosphorylation of TFEB and TFE3. Given the value of p38 MAP kinase invarious conditions including oxidative stress, the findings with one another indicate that SB202190 should not be applied like a particular inhibitor for elucidating the p38 MAP kinase biological functions due to its potential effect on activating autophagy-lysosomal axis.